Enzyme Name: thymine diphosphate(dTDP)-Glucose 4,6-dehydratase (rfbB)
EC Number: 4.2.1.46
PDB ID: 6BI4
Organism: Bacillus anthracis
AA Length: 354
Gene Length: 1065
Involved in three pathways: (works with a group of proteins)
Streptomycin biosynthesis
Polyketide sugar unit biosynthesis
Biosynthesis of vancomycin group antibiotics
Ligands on the X-ray structure are sucrose, a sulfate ion, NAD, and a nickel (II) ion.
Substrate is dTDP-d-glucose
dTDP-Glucose 4,6-dehydratase is involved in a pathway that synthesizes rhamnose sugar, a polysaccharide that is incredibly important in cell wall development of Gram-positive bacteria. The role of rhamnose in synthesis of the cell wall was studied in streptococcus and found to comprise 50-60% of the cell wall thickness, and that bacteria that did not produce this sugar had severe growth and cell division abnormalities. This could have the same effect in Bacillus anthracis.
Link to PDB page: http://www.rcsb.org/structure/6BI4
dTDP-glucose 4,6-dehydratase is not similar to any human proteins or even any proteins belonging to any organism in Kingdom Animalia.
There are only a few compounds currently under study as inhibitors of dTDP-glucose 4,6-dehydratase. This means it would be a good enzyme to study because it is not being overstudied as a potential target for inhibiting Bacillus anthracis.
dTDP-glucose 4,6-dehydratase is highly expressed in the organism because it plays a large role in the synthesis of the cell wall of Bacillus anthracis. Based on experiments that have already been completed, dTDP-glucose 4,6-dehydratase can be expressed in E. Coli using a plasmid called rfbB.
In the majority of past completed assays, the plasmid was created from scratch using PCR. This plasmid was then transformed into E. coli and expressed.
Gene sequence:
Atgaaattcctcattaccggcggagccggctttataggctccgcagttatccgccacatcatcaaccacactcaagacagcgtggtcaaccttgataagttgacctacgcaggcaatcttgaatctttggcttcggttgcagacaacccgcgttatacatttgaacaggcagatatttgcgaccgctctgcgcttgagtggattttcttgcaccatcaaccggatgccgttatgcacttggccgcggaaagccatgtcgaccgctcgatcgacgggccggcggcgtttattgaaaccaatatcgtcggcacttataccttgttggaggcagcacgggcatactggcaaaacatgccgtctgaaaaacaggcgtcgttccgttttcatcatatttctaccgacgaagtgtatggcgatttgcacggtaccgacgatttgtttaccgaaaccacgccttatgcgcccagcagcccttattccgcttccaaagcctccagcgaccatttagtaagggcatggcggcgcacttacggcttgccttgtttgattaccaattgttccaacaactacggcccttatcattttcctgaaaaactgattcctttgatgattctcaatgccatcggcggcagacccttgcccgtttacggcaacggtttgcagatacgcgattggctgtttgtcgaagatcatgcgcgggctttatatgccgtgatcacacaggggaaaataggggaaacctataatatcggcggccataacgaaaaaaccaatctcgaagtggtgcaaaccgtctgcaaactgctggaagaattgatgcccgaaaaacccgccggcgttcgagcctatgaagatttgatcacgtttgtcgaagaccgccccggccacgatgtgcgttatgccattgatgcggccaaaatcaaccacgagcttggctggaagcccgaagaaacttttgaatcgggtatccgtaaaaccgtagcatggtatttgaacaaccgcggctggtggcaacgtgtgctcgacggctcctaccggctcgaacgcttgggagtggcaaaatga
Amino Acid Sequence:
MKFLITGGAGFIGSAVIRHIINHTQDSVVNLDKLTYAGNLESLASVADNPRYTFEQADICDRSALEWIFLHHQPDAVMHLAAESHVDRSIDGPAAFIETNIVGTYTLLEAARAYWQNMPSEKQASFRFHHISTDEVYGDLHGTDDLFTETTPYAPSSPYSASKASSDHLVRAWRRTYGLPCLITNCSNNYGPYHFPEKLIPLMILNAIGGRPLPVYGNGLQIRDWLFVEDHARALYAVITQGKIGETYNIGGHNEKTNLEVVQTVCKLLEELMPEKPAGVRAYEDLITFVEDRPGHDVRYAIDAAKINHELGWKPEETFESGIRKTVAWYLNNRGWWQRVLDGSYRLERLGVAK
Assay Information:
Substrates are readily available for purchase from Sigma.
Absorbance assay at 320 nm using a microplate spectrophotometer.
Description: Macintosh HD:Users:taylorgardiner:Desktop:6bi4.pdb1-500.jpg
EC Number: 4.2.1.46
PDB ID: 6BI4
Organism: Bacillus anthracis
AA Length: 354
Gene Length: 1065
Involved in three pathways: (works with a group of proteins)
- Streptomycin biosynthesis
- Polyketide sugar unit biosynthesis
- Biosynthesis of vancomycin group antibiotics
Ligands on the X-ray structure are sucrose, a sulfate ion, NAD, and a nickel (II) ion.Substrate is dTDP-d-glucose
dTDP-Glucose 4,6-dehydratase is involved in a pathway that synthesizes rhamnose sugar, a polysaccharide that is incredibly important in cell wall development of Gram-positive bacteria. The role of rhamnose in synthesis of the cell wall was studied in streptococcus and found to comprise 50-60% of the cell wall thickness, and that bacteria that did not produce this sugar had severe growth and cell division abnormalities. This could have the same effect in Bacillus anthracis.
Link to PDB page: http://www.rcsb.org/structure/6BI4
dTDP-glucose 4,6-dehydratase is not similar to any human proteins or even any proteins belonging to any organism in Kingdom Animalia.
There are only a few compounds currently under study as inhibitors of dTDP-glucose 4,6-dehydratase. This means it would be a good enzyme to study because it is not being overstudied as a potential target for inhibiting Bacillus anthracis.
dTDP-glucose 4,6-dehydratase is highly expressed in the organism because it plays a large role in the synthesis of the cell wall of Bacillus anthracis. Based on experiments that have already been completed, dTDP-glucose 4,6-dehydratase can be expressed in E. Coli using a plasmid called rfbB.
In the majority of past completed assays, the plasmid was created from scratch using PCR. This plasmid was then transformed into E. coli and expressed.
Gene sequence:
Atgaaattcctcattaccggcggagccggctttataggctccgcagttatccgccacatcatcaaccacactcaagacagcgtggtcaaccttgataagttgacctacgcaggcaatcttgaatctttggcttcggttgcagacaacccgcgttatacatttgaacaggcagatatttgcgaccgctctgcgcttgagtggattttcttgcaccatcaaccggatgccgttatgcacttggccgcggaaagccatgtcgaccgctcgatcgacgggccggcggcgtttattgaaaccaatatcgtcggcacttataccttgttggaggcagcacgggcatactggcaaaacatgccgtctgaaaaacaggcgtcgttccgttttcatcatatttctaccgacgaagtgtatggcgatttgcacggtaccgacgatttgtttaccgaaaccacgccttatgcgcccagcagcccttattccgcttccaaagcctccagcgaccatttagtaagggcatggcggcgcacttacggcttgccttgtttgattaccaattgttccaacaactacggcccttatcattttcctgaaaaactgattcctttgatgattctcaatgccatcggcggcagacccttgcccgtttacggcaacggtttgcagatacgcgattggctgtttgtcgaagatcatgcgcgggctttatatgccgtgatcacacaggggaaaataggggaaacctataatatcggcggccataacgaaaaaaccaatctcgaagtggtgcaaaccgtctgcaaactgctggaagaattgatgcccgaaaaacccgccggcgttcgagcctatgaagatttgatcacgtttgtcgaagaccgccccggccacgatgtgcgttatgccattgatgcggccaaaatcaaccacgagcttggctggaagcccgaagaaacttttgaatcgggtatccgtaaaaccgtagcatggtatttgaacaaccgcggctggtggcaacgtgtgctcgacggctcctaccggctcgaacgcttgggagtggcaaaatga
Amino Acid Sequence:
MKFLITGGAGFIGSAVIRHIINHTQDSVVNLDKLTYAGNLESLASVADNPRYTFEQADICDRSALEWIFLHHQPDAVMHLAAESHVDRSIDGPAAFIETNIVGTYTLLEAARAYWQNMPSEKQASFRFHHISTDEVYGDLHGTDDLFTETTPYAPSSPYSASKASSDHLVRAWRRTYGLPCLITNCSNNYGPYHFPEKLIPLMILNAIGGRPLPVYGNGLQIRDWLFVEDHARALYAVITQGKIGETYNIGGHNEKTNLEVVQTVCKLLEELMPEKPAGVRAYEDLITFVEDRPGHDVRYAIDAAKINHELGWKPEETFESGIRKTVAWYLNNRGWWQRVLDGSYRLERLGVAK
Assay Information:
Substrates are readily available for purchase from Sigma.
Absorbance assay at 320 nm using a microplate spectrophotometer.